Modern Delivery Platforms for DNA Cancer vaccines: From Research to Clinical Development

Cancer remains one of the major causes of mortality worldwide, with common types including breast, lung, and oral cancers. The increasing burden of cancer highlights the urgent need for targeted and effective therapeutic strategies. DNA cancer vaccines have emerged as a promising immunotherapeutic approach because of their ability to generate tumor-specific antigens and stimulate immune responses. Conventional treatment methods such as surgery, chemotherapy, and radiotherapy are still widely used; however, these approaches often have limitations, including toxicity and limited long-term effectiveness.

DNA cancer vaccines aim to activate both humoral and cellular immune responses, particularly cytotoxic T-cell immunity, to control tumor growth. However, one of the major challenges in preclinical and clinical studies is the efficient delivery of DNA vaccines. Improper delivery can reduce vaccine effectiveness and limit immune activation.

Skin-based delivery has gained attention as an effective method for DNA vaccine administration. The skin is a highly immunologically active organ containing a large number of antigen-presenting cells (APCs), including dendritic cells and Langerhans cells. The epidermis layer (approximately 0.05–0.2 mm thick) contains melanocytes, Merkel cells, Langerhans cells, and keratinocytes. The underlying dermis layer (approximately 1.5–3 mm thick) contains dendritic cells, mast cells, macrophages, and collagen fibers. These immune cells play an important role in antigen recognition and immune activation.

Therefore, delivering DNA vaccines to the epidermal or dermal layers rather than subcutaneous tissue may improve immune responses. In recent years, several tumor-associated antigens have been identified as potential targets for immunotherapy. Positive immune responses observed in studies indicate the potential effectiveness of DNA vaccines.

Various physical delivery methods such as DNA tattooing, gene gun, ultrasound-mediated delivery, and electroporation have been developed to improve DNA uptake. In addition, these delivery techniques are often combined with chemical formulations, liposomes, or micro- and nanoparticles to enhance targeting and improve vaccine efficiency.

2.DNA vaccine:

DNA plasmid vaccines are generally composed of two main components: a transcriptional unit and an immune-stimulatory unit. The transcriptional unit includes the promoter region responsible for initiating gene expression and the structural gene encoding the antigen. The immune-stimulatory component enhances the activation of the host immune system. DNA cancer vaccines function by encoding tumor-specific antigens, similar to other DNA vaccines, and stimulate immune responses against cancer cells. These vaccines combine principles from immunology, molecular genetics, and personalized medicine, making them an important part of modern cancer immunotherapy.

DNA vaccines are designed to activate CD8? cytotoxic T lymphocytes (CTLs), which play a critical role in tumor destruction. In addition, they can induce both humoral and cellular immune responses. Enhanced antigen presentation by dendritic cells and improved interaction between dendritic cells and T cells further strengthen immune activation. However, one of the major challenges in DNA vaccine development is the efficient delivery of plasmid DNA during preclinical and clinical studies. The effectiveness of DNA cancer vaccines largely depends on selecting appropriate delivery systems.

Various physical delivery methods such as DNA tattooing, gene gun, ultrasound-mediated delivery, electroporation, and contact-independent helium plasma have been investigated to improve DNA uptake. These delivery strategies are often combined with chemical formulations, micro- or nanoparticles to enhance targeting of antigen-presenting cells (APCs). Biological delivery systems typically include viral vectors, while non-biological systems include polymers, liposomes, and cationic peptides. In recent years, several tumor-associated antigens (TAAs) have been identified as promising targets for cancer therapy, and studies have demonstrated favorable immune responses using these targets.

Initial findings from preclinical and clinical studies suggest that optimized delivery methods can significantly enhance DNA vaccine effectiveness. DNA vaccines often require high doses due to limited cellular uptake; however, increasing dosage does not always improve efficacy. Physical and chemical delivery methods can temporarily increase cell membrane permeability, allowing lower doses of DNA vaccines to achieve better results. Therefore, continued research is necessary to optimize biological, non-biological, and physical delivery systems to improve the clinical success of DNA cancer vaccines.

How does the DNA cancer vaccine work

1.Tumor-specific antigens are identified on vancer cells.

2.DNA encoding these antigens is administered into the body.

3.Host cells produce protiens related to tumor antigens.

4.The immune system recognizes these proteins as foreign substances.

5.Immune cells, particularly T lymphocytes, attack cancer expressing these antigens.

Clin Exp Vaccine Res 2024;13:73-82

https://doi.org/10.7774/cevr.2024.13.2.7enable the immune response. These vaccines are depends on DNA encoding tumor associated antigen (TTAs).

Figure.2.1: Comparison of intramuscular (IM) injection, subcutaneous (SC) injection,               and microneedle patch.¹

3.Pre-clinical studies:

Pre-clinical studies have demonstrated encouraging results for DNA cancer vaccines in different animal models. These investigations mainly focus on evaluating safety, immunogenicity, and therapeutic efficacy before clinical testing. DNA vaccines have shown the ability to generate antigen-specific immune responses, including activation of cytotoxic T lymphocytes and antibody production, which are essential tumor elimination.

Animal studies have also indicated that combining DNA vaccines with adjuvants or advanced delivery techniques can enhance immune responses. Among various delivery methods, electroporation has demonstrated improved antigen expression and stronger immune activation compared to conventional injection techniques. These findings highlight the importance of optimizing delivery strategies to improve vaccine effectiveness.

Furthermore, several tumor-associated antigens such as HER2, PSA, and MAGE have been evaluated in pre-clinical studies. These antigens are commonly expressed in different cancers and serve as potential targets for vaccine-based therapy. The promising outcomes from pre-clinical research support further clinical development of DNA cancer vaccines.

Table 3.1: DNA cancer vaccine in clinical trials

4.Delivery systems for DNA cancer vaccines

Efficient delivery of plasmid DNA remains a significant challenge in the development of DNA vaccines. To improve cellular uptake and enhance immune responses, several delivery strategies have been explored. These approaches are broadly classified into physical, biological, and non-biological delivery systems.

4.1 Physical delivery system

Physical delivery approaches utilize mechanical or electrical techniques to introduce DNA into host cells. These methods temporarily increase cell membrane permeability, enabling plasmid DNA to enter cells more effectively and enhance transfection efficiency.

Electroporation

Electroporation involves applying short electrical pulses to create transient pores in the cell membrane. This process facilitates DNA uptake and improves antigen expression. Several studies have shown that electroporation produces stronger immune responses compared with traditional injection techniques.

Gene Gun

The gene gun method delivers DNA-coated gold particles into the skin using high-pressure propulsion. This approach directly targets antigen-presenting cells and promotes immune activation. Gene gun delivery has demonstrated encouraging outcomes in both preclinical and early clinical investigations.

DNA Tattooing

DNA tattooing involves intradermal delivery of plasmid DNA using tattoo-based devices. This technique enhances immune responses by activating skin-resident immune cells and improving antigen presentation.

4.2 Biological delivery systems

Biological delivery systems employ microorganisms such as viruses and bacteria to transport plasmid DNA into host cells. These approaches are extensively studied due to their high transfection efficiency and their ability to induce strong immune responses.

Viral Vectors

Viral vectors, including adenoviruses, retroviruses, and vaccinia viruses, are frequently used for DNA vaccine delivery. These vectors effectively transfer genetic material into host cells, resulting in enhanced antigen expression. However, viral delivery methods may present safety concerns, including immune reactions, toxicity, and the possibility of integration into the host genome.

Bacterial Vectors

Bacterial carriers, such as attenuated strains of Salmonella and Listeria, have also been explored as delivery vehicles. These systems can stimulate immune responses and improve antigen presentation. Despite their potential, additional research is necessary to establish their safety and efficacy in clinical applications.

4.3 Non-Biological Delivery Systems

Non-biological delivery systems include lipid-based carriers, nanoparticles, and polymer-based approaches. These methods enhance DNA stability and promote cellular uptake without relying on biological organisms.

Liposomes

Liposomes are lipid-based vesicles that encapsulate plasmid DNA and assist in delivering it into host cells. These carriers protect DNA from degradation and improve the overall effectiveness of DNA vaccines.

Nanoparticles

Nanoparticles enable targeted delivery and controlled release of DNA vaccines. They enhance immune responses while minimizing toxicity. Various materials, including gold nanoparticles, polymeric nanoparticles, and lipid nanoparticles, have been investigated for this purpose.

Polymer-Based Systems

Polymer-based carriers, such as polyethyleneimine (PEI), improve DNA delivery by increasing cellular uptake and protecting DNA from degradation. These systems are widely studied because of their stability and relatively favorable safety profile

4.4 DNA tattooing

DNA tattooing is an intradermal delivery method that uses tattoo devices to introduce plasmid DNA into the skin. This technique stimulates immune responses by activating antigen-presenting cells present in the skin. Clinical studies have demonstrated that DNA tattooing is safe and capable of inducing strong immune responses. This method is gaining attention as an effective and minimally invasive delivery approach

4.5 Ultrasound

Ultrasound-mediated delivery is a non-invasive technique used to enhance DNA vaccine uptake. This method temporarily increases cell membrane permeability through a process known as sonoporation. Acoustic energy generated by ultrasound creates temporary pores in the cell membrane, allowing plasmid DNA to enter cells.

Preclinical studies have shown that ultrasound delivery can significantly improve immune responses compared to conventional injection methods. Additionally, this technique activates immune cells such as Langerhans cells, further enhancing immune activation.

4.6 Contact-independent helium plasma

Contact-independent helium plasma is a novel, non-invasive delivery technique that uses ionized helium gas to enhance DNA uptake. This method generates charged particles that temporarily increase cell membrane permeability, allowing plasmid DNA to enter cells.

This approach is considered a potential alternative to electroporation, as it reduces discomfort and eliminates the need for needle-based delivery. Studies have shown that helium plasma delivery increases CD8+ cytotoxic T cell responses, which play an important role in tumor destruction

4.7 Electroporation

Electroporation is one of the most effective methods for DNA vaccine delivery. This technique uses electrical pulses to create temporary pores in cell membranes, allowing plasmid DNA to enter cells efficiently. Electroporation enhances antigen expression and improves both cellular and humoral immune responses.

Studies have shown that electroporation can increase DNA uptake by 100 to 1000 times compared to conventional injection methods. This technique has been widely studied in both preclinical and clinical trials for DNA vaccine delivery.

5.Clinical Trials:

Several clinical trials have investigated DNA cancer vaccines in patients with various types of cancer. These studies mainly evaluate safety, immunogenicity, and therapeutic efficacy. Early-phase clinical trials indicate that DNA vaccines are generally safe and well tolerated, with only minimal adverse effects reported.

Although immune responses have been observed in many studies, therapeutic outcomes differ depending on cancer type, antigen selection, and delivery strategy. To enhance treatment effectiveness, researchers are exploring combination therapies that include DNA vaccines with immune checkpoint inhibitors, chemotherapy, and other immunotherapeutic approaches. Recent studies conducted between 2024 and 2025 have reported promising outcomes, particularly when DNA vaccines are used as part of combination treatments.

1. Prostate Cancer DNA Vaccine

Recent clinical trials have evaluated DNA vaccines for prostate cancer management. These studies suggest that DNA vaccines may help delay disease progression rather than produce rapid tumor shrinkage.

pTVG?HP + Nivolumab (Phase II)

Clinical trials involving patients with non-metastatic prostate cancer demonstrated that treatment with pTVG-HP combined with nivolumab delayed disease progression. Although significant decreases in prostate-specific antigen (PSA) levels were not observed, immune activation was detected.

Method:

Patients received pTVG-HP and nivolumab every two weeks for three months, followed by administration every four weeks for one year.

Immune Response:

Antigen-specific T-cell responses were observed in approximately 79% of participants.

pTVG-HP Long-Term Survival

Phase II clinical trial findings showed improved overall survival. Patients treated with the vaccine demonstrated an average survival of 13.4 years compared with 8.6 years in the placebo group.

ADXS31?142 + Pembrolizumab

Phase I/II clinical trials in metastatic castration-resistant prostate cancer reported improved survival outcomes with combination therapy. The combined treatment resulted in survival of approximately 33.7 months compared with 7.5 months for vaccine therapy alone.

Figure 5.1. Tumor Infiltration of therapeutic vaccine.²³

2.Melanoma DNA and RNA vaccine trials

 Recent developments in DNA and mRNA vaccine technologies have demonstrated encouraging outcomes in melanoma treatment.

NeoVaxMI (Phase I)

NeoVaxMI is a personalized DNA vaccine platform evaluated in patients with high-risk melanoma. Findings reported in 2025 indicated favorable safety profiles along with strong immune activation.

VB10.NEO + Pembrolizumab

The VB10.NEO DNA vaccine platform is currently being studied in Phase II clinical trials. This vaccine targets tumor-specific neoantigens and enhances immune responses by delivering DNA directly to antigen-presenting cells.

Comparison with mRNA?4157 Vaccines

Clinical studies combining mRNA-4157 with pembrolizumab reported nearly a 49% reduction in disease progression or mortality. These findings emphasize the growing potential of nucleic acid-based vaccines in cancer immunotherapy.

Figure 5.2 Mechanisms of action of various vaccines in melanoma immunotherapy.²⁴